Mechanisms of heart rate regulation

Question 1

What can heart rate predict?

Answer 1

CVD mortality in acute and chronic disease

• resting HR above 70 beat/min considered to  risk if you have CVD

Question 2

Why is heart rate a predictor of CVD mortality?

Answer 2

• increased HR linked to atherosclerosis/coronary artery plaque disruption and thus may lead to thrombus and occlusion of coronary artery
• HR is determinant of myocardial O2 consumption – high HR implies the heart is less efficient
• Determinants of coronary circulation perfusion time – every time there is a systolic contraction, there’s a reduction in coronary circulation perfusion time, high HR means more time is systole/less time in diastole and there’s a reduction in coronary perfusion

Question 3

Why is decreasing heart rate a target for CVD treatment?

Answer 3

• decreased HR leads to a decrease in O2 demands of heart
• increase in Blood flow to heart
• decrease in HR is a target for treating post-MI, angina, heart failure etc.
• Use of beta1 blockers, Ca2+ channel blockers

Question 4

Where is HR initiated and regulated?

Answer 4

Sino- atrial node (SAN)
• Primary area generating pacemaker potentials in the heart
• Provides the initial electrical stimulus for myogenic activity of the heart
• Direct relationship between pacemaker frequency and heart rate (HR)

Question 5

What is the area of SAN determined by?

Answer 5

• Measuring electrical activity: area affected by vagal stimulation – vagus nerve innervates the SAN
• Staining: can stain neurofilament (SAN + atrial myocytes), Cx43 aka connexin 43 (atrial myocytes), ANP (atrial myocytes release this) – area of no Cx43/ANP but neurofilament staining = SAN

Question 6

What makes SAN cells different from other cells?

Answer 6

electrical activity generating but do not contractile or conduct

Question 7

How do SAN cells generate electrical activity?

Answer 7

Express HCN4 proteins – make up If channels (HCN4 proteins are not present in other areas of the heart), these channels are important for producing electrical activity

Question 8

What are central SAN area surrounded by?

Answer 8

fibrosis/connective tissue:

• SAN cells do not express connexins (e.g. Cx43, like atrial myocytes), has poor gap junction structure
• This means SAN is electrically isolated from rest of heart.

Question 9

Why is it important that SAN cells are electrically isolated from the rest of the heart?

Answer 9

• Pacemaker potentials thought to leave SAN and spread to atrial through controlled specific pathways – currently unclear
• Because it’s isolated SAN is not influenced by atrial electrical activity
• Very important as atrial repolarisations would ‘switch-off’ SAN

Question 10

How does the SAN generated pacemaker potential cause ventricular contraction?

Answer 10

• SAN generated pacemaker potential and other action potentials generated by the electrical activity conducts out of the SAN into the surrounding atrial tissue.
• Potential slows down as it goes into the atrio-ventricular node and it speeds up again as it passes through the bundle of His, the left and right bundle branches, purkinje fibres and ventricular muscle.
• It causes a contraction in the ventricular tissue.

Question 11

How does the SAN generated pacemaker potential make ECG patterns?

Answer 11

The co-ordinated stimulation and repolarisation of action potentials is what causes the ECG pattern.

Question 12

what forms ionic basis for initiating pacemaker activity in the absence of external stimuli?

Answer 12

• Activation of If channels initiates a diastolic depolarisation which forms the ionic basis for initiating pacemaker activity
• There’s an unstable resting membrane potential – keeps generation action potentials

Question 13

What happens in Phase 0?

Answer 13

Phase 0 is the activation of the upstroke – due to activation of voltage-gated Ca channels because Ca comes in, positively charged and causes an upstroke in the action potential

Question 14

What happens in phase 3?

Answer 14

Phase 3 Ca channels switch off and K channels activate, and K moves out of the cell, down it’s concentration gradient, negative charge inside the cell – repolarisation

Question 15

What happens in phase 4?

Answer 15

Phase 4 is the unstable resting membrane potential, If channels are activated.

If channels are hyperpolarisation activated non-selective channels - as the cell hyperpolarises and repolarises the If channels switch on and bring in Na

• the cell becomes positive, starts depolarisation and that continues until threshold is reached for activation of voltage gated calcium channels

Question 16

What do the different phases correlate with?

Answer 16

Different phases of the pacemaker potential relate to different phases of the heart – systole and diastole

Question 17

What does the voltage clock interact with?

Answer 17

The calcium clock

Question 18

What is the calcium clock?

Answer 18

• rhythmic spontaneous sarcoplasmic reticulum Ca2+ release, feeds into voltage clock to cause initiation of action potential
• Sarcoplasmic reticulum releases Ca through RyR, the calcium does several things:
  • Can be taken back up by SERCA
  • NaCa exchanger exchanges it for 3 Na, it gets taken out. The cell become more positive and causes depolarisation.

Calcium also comes in from L-type and t-type calcium channels – causes further activation of the NaCa exchanger and uptake of Ca into store

Question 19

What is the voltage clock?

Answer 19

• cyclic activation and deactivation of membrane ion channels

Question 20

How do the voltage clock and the calcium clock interact?

Answer 20

• A lot of calcium in the sarcoplasmic reticulum of the SAN cells
• SERCA (sarcoplasmic reticulum calcium ATPase) pump uses ATP to take calcium up its concentration gradient, into the sarcoplasmic reticulum
• ryanodine receptors are ligand gated ion channels, allow release of calcium from sarcoplasmic reticulum into cytoplasm
• Voltage clock controlled by calcium channels (bring calcium in). If channels bringing in Na, and potassium channels for repolarisation
• Ca clock feeds into it and initiates voltage clock through the sodium-calcium exchanger

Question 21

What is diastolic depolarisation

Answer 21

Normal cells have a stable resting potential during diastole

- SAN cells have diastolic depolarisation

Question 22

What are the ion channel interactions during diastolic depolarisation?

Answer 22

There’s a lot of important ion channel interactions that occur during diastolic depolarisation

• K channels repolarise, become activated
• If channels: Hyperpolarisation-activated cyclic nucleotide (HCN); activated at

Question 23

What are the phases of diastolic depolarisation?

Answer 23

• Linear: If activation is polarising activating current, causes slow depolarisation in a linear scale
• Non-linear: allows depolarisation threshold to be met earlier and there’s an exponential increase in depolarisation – caused by release of the “tick” Ca from RyR and NaCa exchanger which brings in Na ions which will cause increased depolarisation.

There’s also increased activity of T-type and L-type channel causes the upstroke after diastolic depolarisation

Question 24

Why are Ca signals bigger during upstroke?

Answer 24

Ca signals are bigger during upstroke because they are coming in through the L-type calcium channel

Question 25

Evidence about what comes first – Voltage or Ca2+ Clock?

Answer 25

* There’s an increase in Ca signalling before action potential is generated – happens during the non-linear diastolic depolarisation and this suggests that Ca signal is generating another current (INaCa) which causes the non-linear diastolic depolarisation * After this there is a much larger upstroke – Ca is coming in through the opening of voltage gated Ca channels (l-type) * LCRs are localised Ca2+ releases * Not influenced by depolarisation * Occur during late diastolic depolarisation

Question 26

What determines speed of Ca2+ Clock – its ticks:

Answer 26

* Speed of release/depletion of SR Ca2+ stores – RyR activity * Speed of SR Ca2+ recycling – SR SERCA activity

Question 27

What is the calcium clock influenced by?

Answer 27

Influenced by: • Constitutive PKA activity (affects RyR) o SAN express constitutively active adenylate cyclase isoforms o Produces cAMP-mediated PKA phosphorylation of RyR o Increases opening of RyR and greater release of Ca2+ from SR • Pacemaker potential frequency (affects SERCA) o More Ca2+ influx through T/L-type Ca2+ channels, o Greater uptake of Ca2+ into stores o More to be released

Question 28

Summary of evidence for Ca2+ Clock drives voltage Clock

Answer 28

1) Block of Ca2+ cycling • Buffering [Ca2+]i to low levels slows/stops pacemaker activity 2) Block RyR • reduced LCRs and reduced pacemaker potential frequency 3) Block L-type Ca2+ channels or prevent depolarisation • reduced Ca2+ entry, reduced SR refilling, block LCRs, and pacemaker failure

Question 29

How are ryanodine receptor linked to rise in calcium?

Answer 29

* Ryanodine - RyR inhibitor, stops localised ca release and there’s only linear diastolic depolarisation due to If channels * Action potential still produced but low frequency and takes longer to meet the threshold and produce upstroke * Proves ryanodine receptor is linked to a rise in calcium

Question 30

How is it proved that INCX (sodium calcium exhanger channels) are involved in diastolic depolarisation?

Answer 30

* Li+ is a NCX (sodium calcium exhanger) inhibitor * Li blocks the NCX thus also blocks a current that triggers the non-linear diastolic depolarisation and the pacemaker potential is not made * INCX - involved in exponential increase in diastolic depolarisation

Question 31

What is seen in Cardiac-selective HCN4 KO mice? (they did not have If channels)

Answer 31

* Decreased If currents * HCN4 KO produces bradycardia + death * Decreased Pacemaker Potential freq If B1 (beta 1) receptors are stimulated it mimics the sympathetic system and causes an increase in HR • B1 effect is via Ca Clock or other HCN channels and not If channels

Question 32

How do Sympathetic and Parasympathetic nervous systems alter rate of diastolic depolarisation

Answer 32

Sym - B1 - Gs - increase AC - increase cAMP – increase If – faster rate of diastolic depolarisation - Remember : increased cAMP leads to increased PKA activity, and PKA-phosphorylation of RyR induces more LCRs, evoking increased INCX Parasym - M2 - Gi - decreased AC - decreased cAMP – decreased If – slower rate of diastolic depolarisation, will reduce heart rate Remember: this will also reduce Ca2+ clock

Question 33

What are If channels mediated by?

Answer 33

If channels mediated by HCN proteins channels, they are clinical targets

Question 34

What do If channels do?

Answer 34

If channels – make ‘funny’ currents • Activated by membrane hyperpolarisation, unique in vertebrates (normally voltage-gated channels activated by depolarisation) • HCN are the molecular correlates of If channels • Four distinct members (HCN1-4) • Expressed HCN cDNA in cell lines – you get If channel currents • Increased activity by cAMP can cause it – more If channels activates cause of it

Question 35

Where is HCN4 expressed in the heart and what is the knockout mouse phenotype for it?

Answer 35

- Most found in SAN but also present in AVN (and His-Purkinje fibres) - K/O mouse phenotype: Varying (mild to marked) effects on cardiac automaticity; embryonic lethality

Question 36

How many types of HCN are there?

Answer 36

4

Question 37

what is Ivabradine?

Answer 37

* A HCN modulator * Ivabradine is the only If channel blocker clinically available * Blocks all HCN isoforms * Little effect on other ion channels (Na+, K+, Ca2+) * Blocks If currents * Prolongs pacemaker potentials – reduction in heart rate * Oral, 50% bioavailability, metabolised by cytochrome P450 into active metabolite , half-life is 2 hours, metabolite is 13 hours * decreased heart rate by 10-20 beats/min in healthy individuals – good safety profile

Question 38

What is the clinical evidence for ivabradine?

Answer 38

* INITIATIVE trial - Ivabradine not inferior to atenolol (beta blocker) to treat symptoms of stable angina * ASSOCIATE trial - add-on to beta-blockers, improved exercise duration • SHIFT trial o Class II/III NYHAS heart failure, EF<35%, >70 bpm, sinus rhythm o Results All CV death decreased 1-2 %, Hospitalisations for HF decreased 26% o Death from HF decreased 26% SIGNIFY trial: Study assessInG the morbidity-mortality beNefits of the If inhibitor Ivabradine in patients with coronarY artery disease • End points reached of death from CV disease and non-fatal MI = unchanged • No positive effect – different to SHIFT trial • decreased HR too much, majority given too high dose (10 mg) • Drug interactions with other drugs • Difference in pathology of HF (heart failure) (SHIFT) vs. CAD (coronary artery disease) (SIGNIFY)